We have studied programmed cell death in the nervous system and the biochemical mechanism of apoptosis in general. To approach the nervous system, more sensitive and in situ methods are needed to identify cells undergoing programmed cell death. We have developed two new methods to identify apoptotic cells under the microscope. (1) We have found that thymocyte-programmed cell death can be followed morphologically with Nomarski optics and that the thymocyte death resembles neuronal cell death. The morphologic analysis of nuclear disintegration has allowed us to test whether cell death is due to production of a toxic factor or due to the loss of a protective factor. Using the new microscopic method to identify apoptosis, the nuclei in the heterokaryons were found to follow the original and distinct fate of the parent cells and not to transfer apoptosis nor viability between nuclei. This new method also allowed us to identify apoptosis as the method of cerebellar granule cell death after MPP + treatment in vitro. (2) We have also developed a molecular detection method to measure DNA strand breaks in situ. This allows us to examine brains of animals undergoing neurodegenerative changes during ischemia, MPTP treatment, and during development. This new method has allowed us to determine the role apoptosis plays during development and during various disease states of the nervous system. 3) We have developed monoclonal antibodies against apoptosis regulatory genes Bcl-2, Bcl-x and Bax to probe the regulation of apoptosis in vitro and in vivo.